Decorating the surface of Escherichia coli with bacterial lipoproteins: a comparative analysis of different display systems.

BACKGROUND: The display of recombinant proteins on cell surfaces has a plethora of applications including vaccine development, screening of peptide libraries, whole-cell biocatalysts and biosensor development for diagnostic, industrial or environmental purposes. In the last decades, a wide variety of surface display systems have been developed for the exposure of recombinant proteins on the surface of Escherichia coli, such as autotransporters and outer membrane proteins. RESULTS: In this study, we assess three approaches for the surface display of a panel of heterologous and homologous mature lipoproteins in E. coli: four from Neisseria meningitidis and four from the host strain that are known to be localised in the inner leaflet of the outer membrane. Constructs were made carrying the sequences coding for eight mature lipoproteins, each fused to the delivery portion of three different systems: the autotransporter adhesin involved in diffuse adherence-I (AIDA-I) from enteropathogenic E. coli, the Lpp'OmpA chimaera and a truncated form of the ice nucleation protein (INP), InaK-NC (N-terminal domain fused with C-terminal one) from Pseudomonas syringae. In contrast to what was observed for the INP constructs, when fused to the AIDA-I or Lpp'OmpA, most of the mature lipoproteins were displayed on the bacterial surface both at 37 and 25 °C as demonstrated by FACS analysis, confocal and transmission electron microscopy. CONCLUSIONS: To our knowledge this is the first study that compares surface display systems using a number of passenger proteins. We have shown that the experimental conditions, including the choice of the carrier protein and the growth temperature, play an important role in the translocation of mature lipoproteins onto the bacterial surface. Despite all the optimization steps performed with the InaK-NC anchor motif, surface exposure of the passenger proteins used in this study was not achieved. For our experimental conditions, Lpp'OmpA chimaera has proved to be an efficient surface display system for the homologous passenger proteins although cell lysis and phenotype heterogeneity were observed. Finally, AIDA-I was found to be the best surface display system for mature lipoproteins (especially heterologous ones) in the E. coli host strain with no inhibition of growth and only limited phenotype heterogeneity.

Strategies to Tackle Antimicrobial Resistance: The Example of Escherichia coli and Pseudomonas aeruginosa.

Traditional antimicrobial treatments consist of drugs which target different essential functions in pathogens. Nevertheless, bacteria continue to evolve new mechanisms to evade this drug-mediated killing with surprising speed on the deployment of each new drug and antibiotic worldwide, a phenomenon called antimicrobial resistance (AMR). Nowadays, AMR represents a critical health threat, for which new medical interventions are urgently needed. By 2050, it is estimated that the leading cause of death will be through untreatable AMR pathogens. Although antibiotics remain a first-line treatment, non-antibiotic therapies such as prophylactic vaccines and therapeutic monoclonal antibodies (mAbs) are increasingly interesting alternatives to limit the spread of such antibiotic resistant microorganisms. For the discovery of new vaccines and mAbs, the search for effective antigens that are able to raise protective immune responses is a challenging undertaking. In this context, outer membrane vesicles (OMV) represent a promising approach, as they recapitulate the complete antigen repertoire that occurs on the surface of Gram-negative bacteria. In this review, we present Escherichia coli and Pseudomonas aeruginosa as specific examples of key AMR threats caused by Gram-negative bacteria and we discuss the current status of mAbs and vaccine approaches under development as well as how knowledge on OMV could benefit antigen discovery strategies.

Moraxella catarrhalis evades neutrophil oxidative stress responses providing a safer niche for nontypeable Haemophilus influenzae.

Moraxella catarrhalis and nontypeable Haemophilus influenzae (NTHi) are pathogenic bacteria frequently associated with exacerbation of chronic obstructive pulmonary disease (COPD), whose hallmark is inflammatory oxidative stress. Neutrophils produce reactive oxygen species (ROS) which can boost antimicrobial response by promoting neutrophil extracellular traps (NET) and autophagy. Here, we showed that M. catarrhalis induces less ROS and NET production in differentiated HL-60 cells compared to NTHi. It is also able to actively interfere with these responses in chemically activated cells in a phagocytosis and opsonin-independent and contact-dependent manner, possibly by engaging host immunosuppressive receptors. M. catarrhalis subverts the autophagic pathway of the phagocytic cells and survives intracellularly. It also promotes the survival of NTHi which is otherwise susceptible to the host antimicrobial arsenal. In-depth understanding of the immune evasion strategies exploited by these two human pathogens could suggest medical interventions to tackle COPD and potentially other diseases in which they co-exist.

Corrigendum: Vaccines Against Antimicrobial Resistance.

[This corrects the article on p. 1048 in vol. 11, PMID: 32582169.].

Vaccines Against Antimicrobial Resistance.

In the last century, life expectancy has increased considerably, thanks to the introduction of antibiotics, hygiene and vaccines that have contributed to the cure and prevention of many infectious diseases. The era of antimicrobial therapy started in the nineteenth century with the identification of chemical compounds with antimicrobial properties. However, immediately after the introduction of these novel drugs, microorganisms started to become resistant through different strategies. Although resistance mechanisms were already present before antibiotic introduction, their large-scale use and mis-use have increased the number of resistant microorganisms. Rapid spreading of mobile elements by horizontal gene transfer such as plasmids and integrative conjugative elements (ICE) carrying multiple resistance genes has dramatically increased the worldwide prevalence of relevant multi drug-resistant human pathogens such as Staphylococcus aureus, Neisseria gonorrhoeae, and Enterobacteriaceae. Today, antimicrobial resistance (AMR) remains one of the major global concerns to be addressed and only global efforts could help in finding a solution. In terms of magnitude the economic impact of AMR is estimated to be comparable to that of climate global change in 2030. Although antibiotics continue to be essential to treat such infections, non-antibiotic therapies will play an important role in limiting the increase of antibiotic resistant microorganisms. Among non-antibiotic strategies, vaccines and therapeutic monoclonal antibodies (mAbs) play a strategic role. In this review, we will summarize the evolution and the mechanisms of antibiotic resistance, and the impact of AMR on life expectancy and economics.